US Using Monkeypox As Surrogate/Model For Smallpox
From Patricia Doyle, PhD
dr_p_doyle@hotmail.com
6-13-3

Monkeypox has been the focus of research for many years. In 2000, CSIRO in
Australia, added the IL 4 gene to monkeypox and found that this action caused
monkeypox to become "weaponized" with a 100% kill rate. CSIRO scientists
claimed that the purpose of the experiment was to alter monkeypox for use
as rodent birth control. Although the experiment was not successful in as
much as rodent birth control, the scientists still submitted the "unexpected"
result a"blueprint" for weaponzied poxivirus to medical journals
for all to see.

Ft. Detrick, as well as other US government and private labs have also been
researching monkeypox.

I do hope that more information will be forthcoming about the true origin
of the US monkeypox outbreak. Once that a pathogen has emerged, it is very
difficult to contain an outbreak and to ensure that the pathogen does not
become ingrained and endemic. I think that it is going to be very difficult,
if not impossible, to purge the US of Monkeypox. We did not do well with other
diseases, such as West Nile Virus, HantaVirus especially Sin Nombre, SARS
etc.

The following website, asmbiodefense.org/tuepos.asp has many biodefense research
article/abstracts, even including work on the 1918 influenza.

IV Cidofovir (approved for treatment of cytomegalovirus retinitis in AIDS
patients) is available for contingency use to treat cases of smallpox under
IND,s held by both DHHS and DoD. Evidence supporting IND use is the ability
of cidofovir to inhibit Variola (smallpox) and other orthopoxviruses in vitro
and to treat mice lethally infected with cowpox by either the intranasal or
aerosol routes. Aerosol infection of cynomolgus monkeys with monkeypox produced
a lethal fibrinonecrotic bronchopneumonia, which could be treated with cidofovir
by using a treatment regimen that mimicked human treatment. To determine if
a lesional model could be successfully treated, we used the uniformly lethal
intravenous monkeypox model to show that cidofovir prophylaxis provided complete
protection, showed no signs of illness, and controlled viral replication as
measured in blood, while the placebo-treated animal had >850 lesions and
blood viral titers, measured by quantitative real-time TAQMAN®-MGB PCR,
first detectable on day 3, rising to 107 genomes/ml by day 9 and remaining
at high levels until the primate was enthanized moribund on day 12. Similar
studies of intravenous infection with 108 PFU of the Harper strain of Variola
produced a similar disease with > 250 lesions (WHO category "grave)
and 33% mortality (day 11), but increasing the viral challenge dose 10-fold
resulted in a 100% acutely lethal disease (mean time to death 4 days) that
more closely mimicked hemorrhagic smallpox, with viral levels in organs 1,000-
to 10,000-fold greater than with 108 PFU. Prophylaxis with IV cidofovir resulted
in protection from lethal disease, with significant reductions of blood viral
levels and lesion counts. We believe the lesional model of smallpox produced
by 108 PFU of Harper strain is the most appropriate primate model for drug
evaluation.

Following the level of virus in blood can be a useful tool in disease management,
especially in monitoring the success of antiviral therapy. Primate models
of smallpox and monkeypox have provided specimens appropriate for validating
methodology and determining if viral burden, as reflected by virus levels
in blood, are useful in monitoring successful experimental antiviral therapy
with IV cidofovir. Blood of experimentally infected primates was extracted
with the Qiagen QIAamp DNA Mini Kit. Incubating blood or tissues in AL buffer
plus Proteinase K for > 1 h at 56°C completely inactivated Variola
(the virus that causes smallpox) and monkeypox. Consequently, 4 h incubation
was adopted as adequate to safely remove Variola-infected samples from biocontainment.
Quantitative TAQMAN®-MGB PCR with a pan-orthopox primer/probe set directed
against the viral hemagglutinin gene allowed us to evaluate virus levels of
1000 gene copies per ml of whole blood or gram of tissue. Virus levels in
blood of cynomolgus monkeys experimentally infected with Variola and monkeypox
were significantly reduced in monkeys successfully treated with cidofovir
compared with levels in controls (placebo: monkeypox > 107 and Variola
> 5 x 108 genomes/ml). To understand how the level of virus in blood correlates
with virus levels in tissues, we used the uniformly lethal intranasal infected
cowpox mouse model as a surrogate to correlate virus in blood and tissues
throughout the course of this lethal infection. Virus was first detectable
in blood at 48 h after infection and peaked at greater than 105 genomes per
ml by day 5 postinfection. Near real-time monitoring of virus levels in blood
may allow for rapid evaluation of the efficacy of antiviral therapy during
an initial smallpox outbreak.

Background: Certain cytokines, particularly interferons (IFN), are well known
for their antiviral effects in various virus infections. In this report, we
demonstrate that IL-15, IFN-â and -ã are capable of enhancing
the extrinsic antiviral activity of cultured murine macrophage cells. Furthermore,
we have adapted a mouse respiratory vaccinia infection model to evaluate the
in vivo antiviral effects of cytokines. Methods: RAW 264.7 cells were activated
with individual cytokines and then co-cultured with vaccinia virus-infected
human 293 cells overnight. The virus titers in the cocultures were measured
by plaque assay. Balb/c mice were infected intranasally with 8 X LD50 of vaccinia
virus (strain WR). Murine cytokines were administered intranasally for 5 consecutive
days, starting one day before the infection. The animals were observed for
signs of sickness and death for 21 days. Results: Our results indicate that
IL-15, IFN-â and -ã are capable of inducing the inhibition of
virus replication in by-stander cells. Intranasal administration of recombinant
murine IFN-á and IFN-ã resulted in animal survival rates of
100 and 95%, respectively. In contrast, no mice in untreated control (placebo)
survived the lethal vaccinia challenge. The organ virus titers in treated
mice were 100,000-fold lower than that in placebo. IL-15 provided slight protection
(10% survival). Both innate and adaptive host immune responses are likely
to play significant roles in the IFN-mediated control of poxvirus replication
in mice. Conclusions: Intranasal administration of IFN-á and IFN-ã
provides nearly full protection against respiratory vaccinia virus infection
in mice; therefore, our results have implications in the utilization of interferon
as a prophylactic tool on a scenario of aerosol orthopoxvirus infection.

http://www.biomedcentral.com/news/20030613/04

June 13, 2003 Previous

Monkeypox in the news
Scientists at US agencies have been using virus as surrogate and model for
smallpox | By Peg Brickley

Long before it became the topic of hourly news bulletins, monkeypox was a
virus of interest to US researchers intent on creating safer smallpox vaccines
and treatments. Some hope now that monkeypox's Western Hemisphere debut will
spread the wealth of scientific knowledge back to central and western Africa,
where monkeypox is endemic and sometimes fatal.

"Monkeypox has not been established as a top-priority disease, and it
should be," said Joel Breman, who chaired the World Health Organization's
monkeypox study committee and is now senior scientist at the National Institute
of Health's John E. Fogarty International Center. "It should be a higher
priority because of its potential for exportation, number one, and number
two, because the natural history of monkeypox remains unknown even 30 years
after its discovery in humans and 45 years from its first discovery in nonhuman
primates."

Labs at the US Army Research Institute for Infectious Diseases (USAMRIID)
and at the National Institutes of Health (NIH) labs in Maryland use monkeypox
as a substitute for smallpox because it is easier to handle in the laboratory
and less likely to spread among humans.

"Our main goal has been to look at therapeutics. We use monkeypox because
it is a BSL-3 [biosafety level 3] agent so we can use it under slightly less
strict conditions in the lab," said Robert Baker, a USAMRIID virologist.

Science and logistics make monkeypox a good surrogate for the more lethal
pox. "Monkeypox produces a disease in monkeys and people that is essentially
identical to smallpox," Baker told The Scientist.

Animal models created at USAMRIID are being used to test therapeutics against
smallpox, such as the antiviral cidofovir as well as safer versions of the
smallpox vaccine, such as the modified vaccinia Ankara (MVA) vaccine, that
potentially pose less of a threat to immunocompromised people.

"There are whole groups of people contraindicated for vaccination,"
Baker said. "One of our goals is to find an option for people who shouldn't
be taking the vaccine. In addition, we are trying to extend the window of
protection so we can treat post-exposure."

In collaboration with Donald Smee, research professor at Utah State University's
Institute for Antiviral Research, Army scientists are developing rodent models
to replace the more expensive primate test subjects for monkeypox. Monkeypox's
importation to this country, probably by way of a giant Gambian rat, was "a
total surprise," Smee, who has published extensively on monkeypox, told
The Scientist. And no, Baker said, prairie dogs were never on the list of
candidates to serve as rodent models. Cotton rats show the most promise, Baker
said.

Bernard Moss, chief of the viral disease laboratory at NIH, is evaluating
highly attenuated MVA against primates infected with monkeypox at USAMRIID.
Ongoing work is also looking at recombinant proteins, Moss said, and he expects
to report results before the end of the year.

"The top priority is to control and mute this epidemic so there aren't
more cases," Breman told The Scientist. "This is a huge opportunity
both to look at treatments and possibly prevention, to look at how the disease
will be acquired and spread in an environment where you can actually do surveillance
properly."